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Membrane-Based Oligonucleotide Array Developed from Multiple Markers for the Detection of Many Phytophthora Species

January 2013 , Volume 103 , Number  1
Pages  43 - 54

Wen Chen, Zeinab Robleh Djama, Michael D. Coffey, Frank N. Martin, Guillaume J. Bilodeau, Lorien Radmer, Geoff Denton, and C. André Lévesque

First, second, and eighth authors: Agriculture & Agri-Food Canada, Central Experimental Farm, Ottawa, Ontario K1A 0C6, Canada; third author: Department of Plant Pathology and Microbiology, University of California, Riverside 92521; fourth, fifth, and sixth authors: United States Department of Agriculture–Agricultural Research Service, Salinas, CA 93905; and seventh author: The Royal Horticultural Society, Wisley, Woking, Surrey, UK GU23 6QB.

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Accepted for publication 24 September 2012.

Most Phytophthora spp. are destructive plant pathogens; therefore, effective monitoring and accurate early detection are important means of preventing potential epidemics and outbreaks of diseases. In the current study, a membrane-based oligonucleotide array was developed that can detect Phytophthora spp. reliably using three DNA regions; namely, the internal transcribed spacer (ITS), the 5′ end of cytochrome c oxidase 1 gene (cox1), and the intergenic region between cytochrome c oxidase 2 gene (cox2) and cox1 (cox2-1 spacer). Each sequence data set contained ≈250 sequences representing 98 described and 15 undescribed species of Phytophthora. The array was validated with 143 pure cultures and 35 field samples. Together, nonrejected oligonucleotides from all three markers have the ability to reliably detect 82 described and 8 undescribed Phytophthora spp., including several quarantine or regulated pathogens such as Phytophthora ramorum. Our results showed that a DNA array containing signature oligonucleotides designed from multiple genomic regions provided robustness and redundancy for the detection and differentiation of closely related taxon groups. This array has the potential to be used as a routine diagnostic tool for Phytophthora spp. from complex environmental samples without the need for extensive growth of cultures.

Additional keywords: DNA hybridization.

© Her Majesty the Queen in Right of Canada, as represented by the Minister of Agriculture and Agri-Food Canada, 2013